c0001 diversity and ecology of eukaryotic marine phytoplankton...

CHAPTER ONE

C0001Diversity and Ecology ofEukaryotic Marine PhytoplanktonFabrice Not*,y, Raffaele Sianoz, Wiebe H.C.F. Kooistrax,Nathalie Simon*,y, Daniel Vaulot*,y, and Ian Probert{,1*UPMC University Paris 06, UMR 7144, Station Biologique de Roscoff, 29680 Roscoff, FranceyCNRS, UMR 7144, Station Biologique de Roscoff, 29680 Roscoff, FrancezIFREMER, Centre de Brest, DYNECO/Pelagos, BP 70 29280 Plouzané, FrancexSZN, Stazione Zoologica Anton Dohrn, Villa Comunale, 80121 Naples, Italy{UPMC University Paris 06, FR 2424, Station Biologique de Roscoff, 29680 Roscoff, France1Corresponding½AU1� author:

Contents

1. Phytoplankton Features 21.1. Diversity of Phytoplankton 21.2. Size Matters 41.3. Global Ecological Patterns 51.4. Current Conceptual Challenges 7

2. The Green Phytoplankton: The Chlorophyta 82.1. General Considerations 82.2. The Mamiellophyceae 92.3. Other Prasinophytes 112.4. Trebouxiophyceae 13

3. The Phytoplankton with Calcareous Representatives: The Haptophyta 133.1. Origins of the Haptophyta 133.2. Haptophyte Diversity 153.3. Haptophyte Evolution 173.4. Distribution and Ecology of Haptophytes 18

4. The Multifaceted Phytoplankton: The Dinoflagellates 204.1. Dinoflagellates as Members of the Alveolate Lineage 204.2. Dinoflagellate Diversity 214.3. Dinoflagellate Evolution 244.4. Ecology of Dinoflagellates 26

5. The Siliceous Phytoplankton: The Diatoms 285.1. The Hallmark of the Diatom: The Silica Cell Wall 285.2. Diatom Diversity 295.3. Diatom Life Cycle 325.4. Diatom Evolution 335.5. Diatom Ecology 34

6. Last, But Not Least Relevant: Other Phytoplankton Taxa 356.1. Stramenopiles Other Than Diatoms 35

Advances in Botanical Research, Volume 64 � 2012 Elsevier Ltd.ISSN 0065-2296,http://dx.doi.org/10.1016/B978-0-12-391499-6.00001-3

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6.2. The Cryptophytes 396.3. The Chlorarachniophyta 406.4. The Euglenids 40

7. Concluding Remarks 40References 41

Abstract

Marine phytoplankton, the photosynthetic microorganisms drifting in the illumi-nated waters of our planet, are extremely diverse, being distributed across majoreukaryotic lineages. About 5000 eukaryotic species have been described withtraditional morphological methods, but recent environmental molecular surveysare unveiling an ever-increasing diversity, including entirely new lineages with nodescribed representatives. Eukaryotic marine phytoplankton are significantcontributors to major global processes (such as oxygen production, carbon fixationand CO2 sequestration, nutrient recycling), thereby sustaining the life of most otheraquatic organisms. In modern oceans, the most diverse and ecologically significanteukaryotic phytoplankton taxa are the diatoms, the dinoflagellates, the hapto-phytes and the small prasinophytes, some of which periodically form massiveblooms visible in satellite images. Evidence is now accumulating that manyphytoplankton taxa are actually mixotrophs, exhibiting alternate feeding strategiesdepending on environmental conditions (e.g. grazing on prey or containingsymbiotic organisms), thus blurring the boundary between autotrophs andheterotrophs in the ocean.½TS1�

s0010 1. PHYTOPLANKTON FEATURES

s0015 1.1. Diversity of Phytoplankton

p0010 This chapter provides an overview of current knowledge on the diversityand ecology of the phytoplankton that drift in the illuminated waters of seasand oceans. The term phytoplankton here corresponds to the functionalgrouping of single-celled organisms (prokaryotes and eukaryotes) that havethe capacity to perform oxygenic photosynthesis. Marine phytoplanktonicprokaryotes all belong to the phylum Cyanobacteria within the domainBacteria. In contrast, eukaryotic phytoplankton, the focus of the presentchapter, is taxonomically very diverse, having representatives in all but onelineage of the eukaryotic tree of life (Fig. 1.1). The early evolutionary historyof eukaryotic phytoplankton (and more generally of all plastid bearingeukaryotes) was shaped by series of endosymbiotic events, involving theengulfment of a cyanobacterium by a eukaryote (Chapter II of this volume,De Clerck, Bogaret, & Leliaert, 2012) or the engulfment of a photosyntheticeukaryote by another eukaryote (Chapter III of this volume, Archibald

2 Fabrice Not et al.




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2012). These endosymbiotic events were accompanied by massive genetransfers from the genomes of the endosymbionts to the genome of the host,traces of which can be detected in modern eukaryotic primary producers.

p0015 Historically, the diversity of eukaryotic phytoplankton has been assessedby microscope-based comparison of morphological features. Based on theseobservations, less than 5000 species have been described to date (Simon,Cras, Foulon, & Lemée, 2009; Sournia, Chretiennot-Dinet,& Ricard, 1991;Tett & Barton, 1995), but there is a general agreement that this numberlargely underestimates the real extent of phytoplankton diversity. In the lastdecade, evaluation of environmental diversity using molecular approacheshas highlighted massive undescribed diversity, including whole lineageswithout any cultured representatives and for which only environmental

f0010 Figure 1.1 Schematic phylogenetic tree representing the distribution of phytoplank-tonic taxa across eukaryote lineages (in color). Illustrations of (a) Chlorophyceae, (b)Pseudoscourfieldia sp., (c) Porphyridium cruentum, (d) Gymnochlora dimorpha, (e)Dinoflagellates, (f) Odontella sp. (g) Bolidomonas pacifica, (h) Dictyocha sp., (i) Aur-eococcus anophagefferens, (j) Heterosigma akashiwa, (k) Pinguiochrysis pyriformis, (l)Ochromonas sp., (m) Nannochloropsis salina, (n) Calcidiscus sp., (o) Cryptomonas sp., (p)Euglenids; ‘a, b, e, f, h, j, l, n, o and p’ are adapted from Tomas (1997), ‘c’ adapted fromLee (1999), ‘d’ from Ota, Kudo, and Ishida (2011), ‘g’ adapted from http://tolweb.org/Bolidomonas/142186, ‘i’ from Andersen and Preisig (2000), ‘k’ from Kawachi et al.(2002) and ‘m’ from Van Den Hoek, Mann, and Jahns (1995).

Diversity and Ecology of Eukaryotic Marine Phytoplankton 3




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sequences are available (Massana & Pedros-Alio, 2008; Vaulot, Eikrem,Viprey, & Moreau, 2008). Some of these environmental lineages are sodistantly related to all other groups that they may represent new phyla, oneexample being the picobiliphyte algae (Not et al., 2007). Combination ofmolecular phylogenetic and morphological analyses has repeatedlydemonstrated the existence of cryptic (or pseudocryptic) species, even insupposedly well-known groups such as diatoms and coccolithophores,fuelling the debate concerning species delineation in protists (Amato et al.,2007; Saez et al., 2003). In addition, detailed studies comparing well-definedspecies complexes demonstrate that commonly used molecular markers (e.g.the 18S rRNA gene) often underestimate diversity, particularly for organ-isms like phytoplankton that have huge population sizes and high turnoverrates (Piganeau et al., 2011a). Information on the genetic diversity ofphytoplankton is likely to significantly increase in the future with the adventof environmental meta-barcoding surveys (Bik et al., 2012; Toulza, Blanc-Mathieu, Gourbiere, & Piganeau, 2012). This is particularly true for small-sized phytoplankton for which very few distinguishing morphologicalcharacters are available.

s0020 1.2. Size Matters

p0020 Eukaryotic phytoplankton cells are not only taxonomically very diversebut also span an exceptionally wide size range both between and withintaxonomic groups. Size spectra can even vary temporally and/or spatiallyin response to varying environmental conditions or succession of life cyclestages. Phytoplankton cells span more than three orders of magnitude insize, ranging from picoplankton (0.2–2 mm) up to mesoplankton (0.2–2 mm). Individual cells of most species are solitary, but many species (e.g.most species within the diatom genera Chaetoceros and Thalassiosira, thedinoflagellate Alexandrium catenella, or the haptophyte Phaeocystis) also havethe ability to form chains or colonies. Although exceptions exist, thelargest size classes of marine phytoplankton are generally dominated by‘golden brown’ groups, notably diatoms and dinoflagellates, whereassmallest size classes essentially consist of green algae from the prasinophytelineage (e.g. Ostreococcus tauri, which has a cell diameter less than 1 mm;Chrétiennot-Dinet et al., 1995). In practice, this wide range of cell sizesrequires the deployment of various collecting devices (plankton nets andfiltration on various mesh sizes) and observation methodologies (opticaland electronic microscopy) to characterize phytoplankton diversity. Cell





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size also affects numerous functional characteristics of phytoplankton. Forinstance, because of their large surface to volume ratio which facilitatespassive nutrient uptake, small cells are particularly well adapted to stableand oligotrophic (nutrient poor) waters, whereas larger cells typicallyperform better in mixed and eutrophic (nutrient rich) settings (Finkelet al., 2010; Mara~n�on et al., 2001). Because the marine environmentexhibits heterogeneous physicochemical structures across space and time,cell size is an important feature to consider from an ecological pointof view.

s0025 1.3. Global Ecological Patterns

p0025 Phytoplankton plays a significant role in global ecology and ecosystemfunctioning. First and foremost, phytoplankton species are primaryproducers and contribute to about half of the primary production on theplanet, of which one forth is estimated to occur in oligotrophic waters(essentially performed by the cyanobacteria Prochlorococcus), one forth ineutrophic waters and half in mesotrophic regions (Field, Romari, Gall,Latasa, & Vaulot, 1998). Phytoplankton participates to the global carboncycle through the so-called biological pump, by fixing carbon, a portion ofwhich is subsequently sequestered at depth. Carbon is ultimately buried atthe sea floor for centuries or longer (Falkowski, 2012). Phytoplankton is alsoat the base of virtually all marine food webs. Under specific light andnutrient conditions, some phytoplankton taxa can form large blooms,particularly in coastal waters of temperate seas. Some bloom-formingphytoplankton produce toxins that affect higher trophic levels (harmful algalblooms or HABs), thus having significant ecological and economic impacts(Hinder et al., 2011; Imai & Yamaguchi, 2012). Classically, study of theecology of phytoplankton communities involves one or a combination ofmicroscope-based morphological studies, flow cytometric cell counting,molecular surveys and/or measurement of the presence of specific photo-synthetic pigments (Jeffrey, 1997). Although each approach has its inherentlimitations, general ecological patterns can be drawn from the literature.Eutrophic coastal and continental shelf waters are classically dominated bydiatoms, dinoflagellates and calcifying haptophytes (coccolithophores),groups that contain species that have the capacity to form large blooms,while other groups such as the euglenophytes, cryptophytes and raphido-phytes produce more localized blooms (Assmy & Smetacek, 2009). Openoceans tend to be dominated by groups such as green algal prasinophytes,





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Chrysochromulina-like haptophytes and small stramenopiles like pelagophytesand chrysophytes (Not et al., 2008; Reynolds, 2006).

p0030 Since phytoplankton are primary producers living in a dispersive envi-ronment, abiotic physicochemical factors exert a strong control on thecomposition and dynamics of phytoplankton communities. Several bloom-forming phytoplankton taxa have the ability to bio-mineralize silica orcalcium, which, among other biogeochemical impacts, drives long-termcarbon sequestration by accentuating sinking to the sea floor after bloomevents. Valuable fossil records exist for these bio-mineralizing taxa and theseare extensively used for paleostratigraphy and paleoclimatology. Bio-mineralization is also probably involved in mechanical defense and probablyexplains, at least in part, why diatoms and coccolithophores are ubiquitous inspring blooms in temperate and boreal systems (Smetacek, 2001).

p0035 Besides the control exerted by the zooplankton, which feed uponphytoplankton, the impact of biotic parameters on the global ecology ofphytoplankton has generally not been studied in great detail. There is nowa growing awareness of the impact of viruses and of parasitic and mutualisticsymbiotic interactions on phytoplankton community structure (Brussaardet al., 2008; Siano et al., 2011) and ultimately on global biogeochemicalcycles (Strom, 2008). We refer to Chapter IX of this volume for a review ongenomic insights into the diversity of algal viruses (Grimsley et al. 2012).

p0040 Each phytoplankton lineage employs diverse trophic strategies. Indeed,although phytoplanktonic organisms are primarily photosynthetic, manyexhibit mixotrophic behavior, feeding on prokaryotes or other smallphytoplankton in addition to conducting photosynthesis. This has been wellcharacterized for certain dinoflagellates (e.g. species within the generaGymnodinium or Amphidinium; Lee, 1999) and haptophytes (e.g. Chrys-ochromulina sp.; Kawachi, Inouye, Maeda, & Chihara, 1991). Phytoplanktoncan also live in symbiosis with larger heterotrophic protists such as fora-minifers or radiolarians and also with metazoans (e.g. in coral reefs; (Weber &Medina, 2012). Recently, several lines of evidence (e.g. stable isotopelabelling) promote the conclusion that such mixotrophic strategies are morefrequent than previously thought in the marine environment (Frias-Lopez,Thompson, Waldbauer, & Chisholm, 2009; Liu et al., 2009; Stoecker,Johnson, De Vargas, & Not, 2009). While exogenous abiotic and bioticfactors exert key controls on phytoplankton growth and mortality, internalfactors such as life cycle traits (D’alelio et al., 2010) or control of cell death(Biddle & Falkowski, 2004) fine-tune the regulation of population dynamics.





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s0030 1.4. Current Conceptual Challenges

p0045 Studies of phytoplankton diversity and ecology, and more generally ofmicrobial ecology and evolution, are driven by a number of major unre-solved conceptual challenges, perhaps the foremost of which is the definitionof what is a species. The species stands as a key concept, a basic unit anda common currency for studies of diversity and ecology in any environment;yet, there is no consensus on how to define a species. Phytoplankton speciesare traditionally defined according to morphological features, but (1)comparisons of morphological and molecular data often provide evidencefor cryptic diversity within ‘morphospecies’ (Amato et al., 2007), (2)morphological traits are prone to change under varying environmentalconditions (Pizay et al., 2009), and (3) the smallest phytoplanktonic cellsusually lack distinctive features (Potter, Lajeunesse, Saunders, & Anderson,1997). As for prokaryotes, the classical biological species concept defined byE. Mayr in 1969 (i.e. members of an interbreeding population reproduc-tively isolated from other such groups and capable of producing fertiledescendants; Mayr, 1969) cannot be applied to most microbial eukaryotesdue to the lack of knowledge on sexual reproduction (Silva, 2008). Otherspecies concepts have been proposed (e.g. ecological, phylogenetic,morphological) (De Queiroz, 2007). While progress has been made towardsthe proposition of a unified species concept (De Queiroz, 2007; Samadi &Barberousse, 2006), operationally applicable non-subjective criteria arelacking to circumscribe phytoplankton, and more generally microbial,species.

p0050 Another major scientific puzzle that has its historical roots in the nine-teenth century (O’malley, 2007) and is currently at the centre of an intensedebate in the field of microbial ecology concerns the conceptual principle of‘everything is everywhere, but the environment selects’, postulating that theabundance of individuals in microbial species is so large that dispersal is neverrestricted by geographical barriers (Finlay, 2002). Intuitively, this might bethought to be particularly true for oceanic phytoplankton, unicellulareukaryotes drifting in a dispersive environment. This statement still struc-tures the ecological and evolutionary understanding of microbial distribu-tion (De Wit & Bouvier, 2006). However, microbial ecology no longerrelies on culture-based studies. With the advent of molecular tools, evidenceis accumulating that tends to show that physicochemical barriers do exist formarine plankton and that species are not globally distributed (Casteleynet al., 2010) and can occupy distinct niches (Foulon et al., 2008). In the near





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future, the use of relevant molecular markers coupled to massive sequencingdepth provided by high throughput technologies will probably allow thisquestion to be fully addressed.

p0055 Finally, another unresolved question is that of the paradox of theplankton formulated by G.E. Hutchinson in 1961, who asked ‘why do somany planktonic species co-exist in a supposedly homogeneous habitat? (i.e.under the competitive exclusion principle of Gause, given the limited rangeof resources required for their growth)’. For specific ecosystems, proposedmechanisms to explain the extreme diversity of phytoplankton includespatial and temporal heterogeneity in physical and biological environmentsat different scales, oscillation and chaos generated by internal and externalcauses and self limitation by toxin-producing phytoplankton. A general andwell-accepted theory to explain environmental plankton diversity is still,however, lacking (Roy & Chattopadhyay, 2007). This question is extremelychallenging in the context of the uncertainties mentioned above concerningspecies delineation and enumeration in natural phytoplankton assemblages.

s0035 2. THE GREEN PHYTOPLANKTON: THE CHLOROPHYTA

s0040 2.1. General Considerations

p0060 The Chlorophyta together with the land plants form the green lineage(Viridiplantae). This group arose after an endosymbiotic event betweena cyanobacterium-related organism and a heterotrophic eukaryote that wasat the origin of the Plantae, also named Archaeplastida, a super group ofeukaryotes that also includes the red algae and glaucophytes (Leliaert,Verbruggen, & Zechman, 2011). The extant Streptophyta include the landplants as well as diverse freshwater algal lineages, while the Chlorophytainclude some freshwater algae and all marine representatives (De Clercket al., 2012). The Chlorophyta and Streptophyta possess the followingcommon unique features: a double membrane bound plastid containingchlorophyll b as the main accessory pigment and starch as well as a uniquestellate structure linking pairs of microtubules in the flagellar base. TheChlorophyta form a strongly supported group in molecular phylogenies andare characterized by unique biochemical and ultrastructural features (Leliaertet al., 2011).

p0065 In marine waters, Chlorophyta are especially important within thesmallest size classes, in particular the picoplankton and nanoplankton, whichare formally defined as cells between 0.2–2 and 2–20 mm, respectively. A





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rather small proportion of the lineage belongs to the Trebouxiophyceae(that are mostly freshwater or terrestrial species), but most of the speciesdescribed from marine isolates (Vaulot et al., 2008) and most 18S rRNAgene sequences recovered from the oceanic environment correspond toprasinophytes. Prasinophytes form a polyphyletic assemblage (Fig. 1.2) withvery few common characters and taxonomists are slowly re-organizing thisgroup by creating new classes for each of the existing clades (Guillou et al.,2004; Marin, 2010, p. 14).

s0045 2.2. The Mamiellophyceae

p0070 The recently defined class Mamiellophyceae (Marin & Melkonian, 2010)encompasses three orders (Mamiellales,Dolichomastigales andMonomastigales).

(A) (B)

2 µm

f0015 Figure 1.2 A) Schematic phylogenetic tree of the green algae and land plants lineagesshowing the relationships among major phytoplanktonic taxa and an estimation oftheir ecological significance. Typical representative of each lineage is indicated inbrackets. The overall ecological significance (illustrated by a five-star ranking) issubjective and has been established based on parameters such as abundance, distri-bution, bloom formation, trophic strategies, toxicity, etc. (color code is 1 blue star ¼having freshwater members, 1 red star ¼ important toxic or harmful species, 1–3 greenstars range ¼ other relevant ecological parameters, no stars means multi-cellular or nomarine species). (B) Illustration of two important prasinophytes belonging to theMamiellophyceae. Top: scanning electron microscopy of the common and abundantMicromonas sp (E. Foulon). Bottom: the smallest photosynthetic eukaryote Ostreococcussp. (D. Vaulot).





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From an ecological point of view, the Mamiellales is the most important order(Fig. 1.2), containing three key genera:Micromonas (Butcher, 1952), with the firstdescribed picoplanktonic species Micromonas pusilla, Ostreococcus (Chrétiennot-Dinet et al., 1995), containing the smallest known photosynthetic algal species(0.8-mm cell diameter),Ostreococcus tauri, and Bathycoccus (Eikrem & Throndsen,1990),with a single scale-bearing coccoid species,Bathycoccus prasinos. These threerelated genera, which share few morphological features, are typical of coastalwaters (Cheung et al. 2010; Collado-Fabri, Ulloa, & Vaulot 2011; Medlin,Metfies,Wiltshire, Mehl, & Valentin, 2006; Not et al. 2004) but can also bloomunder specific conditions in oceanic waters (Treusch et al., 2011) or be dominantin Arctic ecosystems (Lovejoy et al., 2007). Members of these three genera canrelatively easily be isolated into pure laboratory culture, facilitating their adoptionas biological and ecological models. Full genome sequences for the threerepresentative genera cited above are now available (Derelle et al., 2006;Moreauet al., Submitted;Worden et al., 2009), and their analysis has started to reveal genesthat are relevant to studies of ecology and speciation (Piganeau et al., 2011b).

p0075 The single described species of the genus Micromonas, M. pusilla, ischaracterized by naked cells with a short flagellum with a characteristichair-point and is genetically differentiated into at least three (but probablymore) clades (Foulon et al., 2008; Guillou et al., 2004; Slapeta, Lopez-Garcia, & Moreira, 2006). Two of the major clades (A and B; sensu Guillouet al. 2004) are found in coastal waters, while clade C is typically oceanic(Foulon et al., 2008). Within clade B, a specific lineage seems to berestricted to Arctic waters (Lovejoy et al., 2007), where it can completelydominate the picophytoplankton size fraction (Balzano, Marie, Gourvil, &Vaulot, 2012).

p0080 Ostreococcus is characterized by small naked coccoid cells with no specificmorphological features except a very salient starch grain in the pyrenoid (Ralet al., 2004). As in the case of Micromonas, it can be subdivided into at leastfour clades based on phenotypic, genetic and genomic traits (Rodriguezet al., 2005). While clade C is mostly restricted to environments where it wasinitially discovered (coastal lagoons), clade A is typical of surface coastalwaters and clade B appears to be associated with deeper layers of theeuphotic zone, displaying specific photoacclimation strategies (Six et al.,2008). However, analyses of the distribution of the 18S rRNA gene ofOstreococcus clades in the Pacific Ocean as well as in the subtropical andtropical North Atlantic indicate that the ecophysiological parametersinfluencing clade distribution are more complex than irradiance alone, withfactors such as temperature and nutrients also being involved in the control





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of the distribution of ecotypes (Demir-Hilton et al., 2011). Ostreococcus canform localized blooms not only in coastal waters (O’kelly, Sieracki, Thier, &Hobson, 2003) but also in open ocean regions (Treusch et al., 2011). Incertain ecosystems, such as the coastal upwelling off Chile, it is the mostabundant picophytoplankton species (Collado-Fabri et al., 2011).

p0085 The third member of the Mamiellales, B. prasinos, is characterized byspider-like scales covering the cell surface (Eikrem & Throndsen, 1990). Incontrast to the two other genera, there is little evidence as yet for theexistence of distinct clades with the genus Bathycoccus (Guillou et al.,2004). Although initially described from the bottom of the euphotic zone(hence the prefix ‘Bathy’; Eikrem & Throndsen 1990), Bathycoccus istypical of surface coastal waters (Collado-Fabri et al., 2011; Not et al.2004). The analysis of metagenomes obtained from sorted cells fromcoastal and pelagic deep chlorophyll maximum waters suggests that theremay indeed be distinct Bathycoccus ecotypes or species adapted to thesedifferent environments (Monier et al., 2012; Vaulot et al., submitted). Twoother nanoplanktonic genera, Mantionella and Mamiella, also belong toMamiellales. The order Dolichomastigales contains two genera Dolicho-mastix and Crustomastix with nanosized cells possessing two very longflagella. The 18S rRNA gene sequences related to these four genera havebeen found in the Mediterranean Sea (Viprey, Guillou, Férréol, & Vaulot,2008), in the Atlantic, and even associated to deep sediment samples(Marin & Melkonian, 2010), but very little information is available ontheir global distribution and ecology.

s0050 2.3. Other Prasinophytes

p0090 The Pyramimonadales (prasinophyte clade I) encompasses more than 35species (Guiry & Guiry, 2012) within the main genus Pyramimonas, char-acterized by nanosized cells typically possessing four flagella. This order canbe ecologically important in coastal areas (Bergesch, Odebrecht, &Moestrup, 2008) as well as in polar waters (Balzano et al., 2012; Rodriguez,Varela, & Zapata, 2002).

p0095 The Chlorodendrophyceae (prasinophyte clade IV) is a recently estab-lished class (Massjuk, 2006), which contains one major genus, Tetraselmis,with around 30 species (Guiry & Guiry, 2012). Cells possess four equalflagella arranged in two opposite pairs and a theca composed of aggregatedscales. This group does not appear to be ecologically important in marinewaters, although related sequences have been found in the Mediterranean





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Sea (Viprey et al., 2008). Cultured strains are widely used for applicationssuch as aquaculture (Mohammady, 2004).

p0100 The Pycnococcaceae (prasinophyte clade V) contains only two majorspecies: Pseudoscourfieldia marina, a flagellate, and Pycnococcus provasolii,a coccoid cell. The two species share 100% 18S rRNA gene identity andcould actually be the two forms of a single life cycle (Fawley, Yun, & Qin,1999; Guillou et al., 2004). Pycnococcus has been found to be abundant inspecific ecosystems such as the Magellan Straits (Zingone, Sarno, Siano, &Marino, 2011). These species are easily isolated from oceanic waters andsimilar sequences have been found, for example, in the Mediterranean Sea(Viprey et al., 2008), suggesting that this group may be widespread.

p0105 As in the case of the Pycnococcaceae, the order Prasinococcales (prasi-nophyte clade VI) contains only two genera, Prasinoderma and Prasinococcus,both falling in the picoplankton size range and containing in total threespecies (Guiry & Guiry, 2012). All three species produce some kind ofgelatinous matrix, which, in the case of Prasinococcus capsulatus, has beenidentified as consisting of a sulfated and carboxylated polyanionic poly-saccharide named capsulan (Sieburth, Keller, Johnson, & Myklestad, 1999).They are easily isolated from marine waters (Le Gall et al., 2008), but few18S rRNA gene sequences are recovered from planktonic environmentalclone libraries (Viprey et al., 2008), suggesting that they may be associated tospecific marine habitats such as marine particles.

p0110 Prasinophyte clade VII has not yet been formerly described, despite thefact that it contains cultured strains, all of which are picosized and coccoid(Vaulot et al., 2008). It is divided into two well-supported subclades (A andB) and, depending on phylogenetic analyses, can include Picocystis salinarum,a small species found in inland saline lakes (Lewin, Krienitz, Goericke,Takeda, & Hepperle, 2000). The 18S rRNA gene sequences from this cladehave been recovered from moderately oligotrophic areas from the PacificOcean and Mediterranean Sea (Shi, Marie, Jardillier, Scanlan, & Vaulot,2009; Viprey et al., 2008) as well as from coastal waters (Romari & Vaulot,2004).

p0115 In contrast to clade VII, no cultures have yet been isolated from prasi-nophyte clades VIII and IX that were first discovered from 18S rRNA genesequences in the Mediterranean Sea (Viprey et al., 2008). Sequences fromclade IX (but not VIII) have also been found in the very oligotrophic watersof the South East Pacific gyre (Shi, Lep�ere, Scanlan, & Vaulot, 2009). Theseclades appear to be extremely diversified and are probably an importantcomponent of the photosynthetic picoplankton in the central oceanic gyres.





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s0055 2.4. Trebouxiophyceae

p0120 Trebouxiophyceae are mostly terrestrial algae, in particular associated withlichens. However, several genera, including Picochlorum (erected to regroupsalt-tolerant Nannochloris species; Henley et al., 2004), Stichococcus andChlorella, can be isolated from marine waters and have been found inenvironmental 18S rRNA gene clone analyses from coastal waters (Medlinet al., 2006).

s0060 3. THE PHYTOPLANKTON WITH CALCAREOUSREPRESENTATIVES: THE HAPTOPHYTA

s0065 3.1. Origins of the Haptophyta

p0125 The haptophytes are a distinct and almost exclusively photosyntheticprotistan lineage that is widespread and often very abundant in diversemarine settings. Haptophytes are characterized by the presence of a uniqueorganelle called a haptonema (from the Greek hapsis, touch, and nema,thread), which is superficially similar to a flagellum but differs in thearrangement of microtubules and in function, being implicated in attach-ment or capture of prey. The group includes some well-known taxa, such asPhaeocystis, Prymnesium and Chrysochromulina, that form periodic harmful ornuisance blooms in coastal environments, the calcifying species (coccoli-thophore) Emiliania huxleyi that produces massive ‘white-water’ blooms inhigh latitude coastal and shelf ecosystems (Fig. 1.3), and Pavlova lutheri andIsochrysis galbana that are species extensively used as feedstock in aquaculture.E. huxleyi has become a model species, notably for studies of the effects ofocean acidification on coccolithophore calcification (Beaufort et al., 2011;Iglesias-Rodriguez et al., 2008; Riebesell et al., 2000), and is the onlyhaptophyte for which extensive genomic data (including full genomesequences) are currently available.

p0130 The origin and evolutionary affiliations of the Haptophyta remaincontentious. Haptophytes were tentatively grouped within stramenopiles(Cavalier-Smith, 1981) since in both lineages plastids contain chlorophyllsa and c as well as various carotenoids, typically giving them a golden orbrown colour, and the photosynthetic carbohydrate storage product is a b-1,3-linked glucan. Haptophytes possess a network of endoplasmic retic-ulum immediately below the cell membrane that was suggested to behomologous to alveoli of ciliates, amphiesmal vesicles of dinoflagellates,the inner membrane complex of apicomplexans, the periplast of





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cryptophytes and possibly mucosal structures of heterokont algae(Andersen, 2004; Cavalier-Smith, 2002; Daugbjerg & Andersen, 1997),supporting the hypothesis that these lineages form a supergroup termedthe chromalveolates, with plastids originating from a single secondaryendosymbiosis event (Cavalier-Smith, 1999). Data from plastid genes havegenerally supported the monophyly of chromalveolate lineages (e.g. Fast,Kissinger, Roos, & Keeling, 2001; Harper & Keeling, 2003), includingevidence from a lateral gene transfer common to the plastids of hapto-phytes and cryptophytes (Rice & Palmer, 2006). However, the chro-malveolate hypothesis also implies that host nuclear lineages aremonophyletic, which has not been confirmed despite the use of substantialgenetic data sets. Nuclear½AU2� -based phylogenomics have consistently shownthat the heterokonts (Stramenopiles) and Alveolates are closely related,forming a strongly supported group with Rhizaria, together constitutingthe so-called SAR group (Burki et al., 2007). Haptophytes generallybranch together with cryptophytes, picobiliphytes and several heterotro-phic groups (telonemids, centrohelids and katablepharids) in these analyses(e.g. Burki et al., 2009). Based on congruent plastid and nuclear data,haptophytes and cryptophytes were proposed to be a distinct chro-malveolate lineage, the Hacrobia (Okamoto, Chantangsi, Horak, Leander,& Keeling, 2009). A recent phylogenomic study based on alignment of258 genes provided strong support for the hypothesis that haptophytes aresister to the SAR group, possibly together with telonemids and cen-trohelids, but that cryptophytes and katablepharids have a common originand are not related to other hacrobians rather than branching with plants(Burki, Okamoto, & Keeling, 2011).

s0070 3.2. Haptophyte Diversity

p0135 The known diversity of extant haptophytes is relatively low compared toother ecologically predominant microalgal groups, with only ca. 400 extantspecies having been described (Jordan, Cros, & Young, 2004). Most hap-tophytes occur as solitary planktonic cells possessing two smooth flagella (i.e.completely lacking mastigonemes) in addition to the haptonema, but soli-tary non-motile planktonic or benthic cells, pseudofilamentous forms andcolonies also exist. Most described haptophytes fall into the nanoplanktonsize class (cells 2–20 m in diameter), but results of environmental molecularsurveys indicate the existence of numerous taxa of very small (<2–3 mm)undescribed picohaptophytes (Liu et al., 2009; Moon-van der Staay et al.,





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2000). Common ultrastructural characters of the group include the presenceof plastids surrounded by four membranes, chloroplast lamellae consisting ofthree thylakoids without girdle (interconnecting) lamellae, tubular mito-chondria and characteristic distension of Golgi vesicles in which organicscales are produced prior to being exported onto the cell surface viaexocytosis. A number of haptophytes are known to undergo a hap-lodiplontic life cycle, with alternation between haploid and diploid phasesboth capable of independent asexual division, each phase characterized bydistinct scale morphology.

p0140 Both morphological and molecular evidence support the division of theHaptophyta into two classes, the Pavlovophyceae and the Prymnesiophy-ceae (Edvardsen et al., 2000). The likely existence of one or more hapto-phyte lineages that occupy an intermediate phylogenetic position betweenthe two described classes has been revealed by analysis of molecular datafrom environmental surveys in marine (Shi et al., 2009) and freshwater(Shalchian-Tabrizi, Reier-Roberg, Ree, Klaveness, & Brate, 2011; Slapeta,Moreira, & Lopez-Garcia, 2005).

p0145 The Pavlovophyceae contains only 13 described species classified ina single order, the Pavlovales (Fig. 1.3). Structural features common to all ormost members of the Pavlovophyceae that distinguish them from thePrymnesiophyceae include the markedly anisokont (i.e. unequal in length)nature of the flagella and the relatively simple arrangement of microtubularand fibrous roots of the pavlovophycean flagellar-haptonematal basalcomplex (Hori & Green, 1994). The Pavlovophyceae are also known tosynthesise certain specific sterols and conjugates, the pavlovols (Véron,Dauguet, & Billard, 1996; Volkman, Farmer, Barrett, & Sikes, 1997) anda unique photosynthetic pigment signature with unknown xanthophyll andtwo polar chlorophyll c forms (Van Lenning et al., 2003). Intriguingly, somepavlovophytes possess an eyespot (Lee, 1999). The organic scales of Pav-lovophyceae, when present, consist of small dense bodies (‘knob scales’) incontrast to the plate scales of the Prymnesiophyceae. The phylogeneticrelationships between known Pavlovales were recently elucidated, leadingto a taxonomic revision of the group (Bendif et al., 2011).

p0150 The vast majority of the known diversity of haptophytes occurs in thePrymnesiophyceae, which comprises two orders of non-calcifying taxa, thePhaeocystales and the Prymnesiales, together with the coccolithophoresmaking up a monophyletic sublineage (the subclass Calcihaptophycidae)containing four orders (Isochrysidales, Coccolithales, Syracosphaerales andZygodiscales; Fig. 1.3).





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p0155 The Phaeocystales contains a single genus, Phaeocystis, with less than 10non-calcifying species, several of which have complex life histories involvingsolitary cells and colonies. The Prymnesiales contains two families of non-calcifying taxa, the Prymnesiaceae and the Chrysochromulinaceae(Edvardsen et al., 2011), the organic plate scales of which are often relativelyhighly elaborated. The two families contain roughly equivalent numbers ofspecies (30–40), but the Chrysochromulinaceae, which are known to be ableto catch prey using their characteristically long haptonema (Kawachi et al.,1991), appear to contain a massive undescribed diversity (possibly hundredsof genotypes) of extremely small taxa (Liu et al., 2009). One recentlydescribed Prymnesiacean species produces siliceous scales (Yoshida, Noel,Nakayama, Naganuma, & Inouye, 2006).

p0160 The coccolithophores possess an exoskeleton of calcareous plates calledcoccoliths. Calcification occurs intracellularly in Golgi-derived vesicles,using organic base-plate scales as the substrate for crystal nucleation. Twomain types of coccoliths exist: heterococcoliths, formed of a radial array ofcomplex-shaped interlocking crystals units, and holococcoliths, constructedof numerous small, similar sized and simple-shaped calcite elements. Het-erococcolith- and holococcolith-bearing taxa were originally thought to bemorphologically and phylogenetically distinct species, but they are nowknown to be different phenotypes exhibited within the life cycle of coc-colithophore species (typically, diploid life cycle stages bear heterococcolithsand haploid stages bear holococcoliths). Although the underlying structuresof coccoliths are universal (Young, Didymus, Bown, Prins, & Mann, 1992),coccolith morphology is extremely diverse, with several morphologicalcategories recognized (Young et al., 2003). Coccolithophore taxonomy isalmost exclusively based on comparison of coccolith morphology; however,molecular studies have demonstrated significant cryptic genetic diversitywithin several coccolithophore species (Saez et al., 2003). Despite theexistence of numerous theories (such as involvement in intracellular supplyof CO2 for photosynthesis, protection from predators or concentration oflight towards plastids; see Young, 1994), the function of coccoliths remainsunknown.

s0075 3.3. Haptophyte Evolution

p0165 Coccoliths are extremely abundant as microfossils, providing an outstandingtool for biostratigraphic dating and studies of evolution. The earliestappearance of coccoliths in the fossil record (corresponding to the origin of





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the calcareous haptophytes) is dated at ca. 220 Mya and several clearevolutionary transitions through the subsequent evolutionary history ofcoccolithophores have been accurately dated (Bown, 1998). This allowscalibration of multiple nodes on phylogenetic trees and hence molecularclock analysis of the evolution of the group as a whole. A recent molecularclock study based on multigene analysis (nuclear 18S rRNA gene or SSU,28S rRNA gene or LSU and plastid tufA and rbcL genes) estimated that thehaptophytes diverged from other chromists in the Neoproterozoic Era ca.824 Mya (1031–637 Mya) around the time of the onset of the Cryogenian‘snowball Earth’ (Liu, Aris-Brossou, Probert, & De Vargas, 2010). In thesame study, the divergence of the two extant haptophyte classes was esti-mated to have occurred 543 Mya, early in the Cambrian period that wit-nessed the most rapid and widespread diversification of life in Earth’s history.The primary radiation within the Prymnesiophyceae (the divergence ofPhaeocystales from other prymnesiophytes) was estimated to have occurred329 Mya in the Carboniferous period, which was characterized by thepresence of widespread shallow epicontinental seas. The timing of the nexttwo divergences within the Prymnesiophyceae, that of the Prymnesiales andthe primary radiation of the Calcihaptophycideae, both apparently followedimportant Earth system transitions early in the Permian and the Triassic,respectively (Liu et al., 2010). Within the Calcihaptophycidae, extant coc-colithophores appear to have diversified from a few lineages that survivedthe major extinction at the Cretaceous/Tertiary (K/T) boundary ca. 65Mya, whereas non-calcifying haptophytes were not affected by the K/Textinction (Medlin, Saez, & Young, 2008). The adaptation of non-calci-fying haptophytes to eutrophic coastal environments and their ability toswitch nutrition modes from autotrophy to mixotrophy were posited aspossible explanations for their survival during this abrupt global changeevent. Members of the Isochrysidalean family Noelaerhabdaceae havenumerically dominated coccolithophore communities for the last 20 millionyears and continue to do so in modern oceans. The noelaerhadacean Emi-liania huxleyi dominates modern coccolithophore assemblages despite thefact that it is a very young species in geological terms, having originated only220 Kya (Thierstein, Geitzenauer, Molfino, 1977).

s0080 3.4. Distribution and Ecology of Haptophytes

p0170 Haptophytes adopt diverse ecological strategies, being present in open-ocean, shelf, upwelling, coastal, littoral, brackish and freshwater





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environments. In addition to undertaking photosynthesis, many hapto-phytes are known to be capable of using particulate and/or dissolved organicfood sources, the group probably therefore being predominantly mixo-trophic (De Vargas, Aubry, Probert, & Young, 2007). Life cycle transitions,with each phase adapted to distinct ecological niches (Noel, Kawachi, &Inouye, 2004), may also be an integral part of the ecological strategy fordifferent haptophytes.

p0175 Pavlovophytes have been described mostly from cultures isolated fromlittoral, brackish water and in some cases freshwater environments, and theyare common components of near-shore planktonic and benthic microalgalcommunities in widespread locations. Due to the difficulty in identifyingpavlovophytes in the light microscope, it is not clear whether theycommonly occur in open-ocean environments. Phaeocystis is ubiquitousfrom poles to tropics and from coastal to open ocean waters and certainspecies regularly produce extensive blooms, notably in Arctic, Antarctic andNorth Sea waters. Phaeocystis blooms can be detrimental to the growth andreproduction of shellfish and zooplankton, and hemolytic and toxic effectshave been reported on fish (Schoemann et al., 2005). Some Prymnesialesspecies also form periodic blooms in coastal waters, occasionally harmful tofish and other biota. The Prymnesiaceae tend to be restricted to coastalwaters, whereas the Chrysochromulinaceae also thrive in oligotrophic openocean regions (Liu et al., 2009) where mixotrophic nutrition is likely animportant strategy. Through the process of calcification and export of calciteto the deep ocean following cell death, coccolithophores play a key role inthe ‘biological pump’ that contributes significantly to global carbon cycling(Rost & Riebesell, 2004). Massive annual blooms of Emiliania huxleyi intemperate and subpolar coastal and shelf environments are visible in satelliteimages (Fig. 1.3). Among the factors that may contribute to the ecologicalsuccess of E. huxleyi are high affinities for inorganic nutrient uptake(Paasche, 2002) and physiological mechanisms for maintaining growthunder high irradiance (Loebl, Cockshutt, Campbell, & Finkel, 2010; Ragni,Aris, Leonardos, & Geider, 2008). Specific viruses play an important role inregulation of E. huxleyi blooms (Wilson et al., 2002). Although it does notform bloom populations in subtropical and tropical environments, E. huxleyi(and the closely related species Geophyrocapsa oceanica) is widespread andrelatively abundant in these ecosystems. The Coccolithales genus Coccolithusincludes relatively large cells (ca. 20 m) that, together with E. huxleyi,dominate coccolithophore communities in subpolar and temperate regionsof the North Atlantic as well as occurring in western-margin upwelling





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zones around the globe. The Coccolithales, Syracosphaerales and Zygo-discales include species that contribute significantly to communities in warmwater mesotrophic settings, including Calcidiscus, Umbilicosphaera, Syr-acosphaera, Helicosphaera, Discosphaera, Rhabdosphaera and Scyphosphaera. Thegenus Syracosphaera is notably very diverse in mesotrophic to oligotrophiccoccolithophore communities. Umbellosphaera, a genus of uncertain phylo-genetic affinities, is abundant in oligotrophic surface layer communities. Thecoccolithophores Florisphaera and Gladiolithus are important members ofdeep-photic zone communities, but the adaptations that allow them tothrive in conditions of extremely low light are unknown.

s0085 4. THE MULTIFACETED PHYTOPLANKTON:THE DINOFLAGELLATES

s0090 4.1. Dinoflagellates as Members of the AlveolateLineage

p0180 The Alveolata constitutes a diverse group of single-celled eukaryotes presentin both marine and terrestrial ecosystems, the principal shared morphologicalfeature of which is the presence of flattened vesicles (cortical alveoli) packedinto a continuous layer supporting the cell membrane (Cavalier-Smith &Chao, 2004). These structures have been associated by immunolocalization toa family of proteins, named alveolins, common to all alveolates (Gould,Tham, Cowman, Mcfadden, & Waller, 2008). Alveolates exhibit extremelydiverse trophic strategies, including predation, photo-autotrophy and intra-cellular parasitism. Most alveolates fall into one of three main subgroups (orphyla): ciliates, dinoflagellates and Apicomplexa, all sharing a commonancestor (Leander & Keeling, 2003). Apicomplexans are obligate parasites ofanimal cells, including humans (e.g. Plasmodinium which causes malaria).Ciliates are mainly aquatic predators that perform essential roles as consumersin microbial food webs, although some taxa can be parasitic or may containsequestered plastids (e.g. Myrionecta rubra). Dinoflagellates are either photo-autotrophs, free-living predators (heterotrophs) or both, either simultaneouslyor alternatively (mixotrophs), while some also live as parasites or symbionts(Taylor, Hoppenrath, & Saldarriaga, 2008). Other lineages of marine alveo-lates (MALV) have been identified in culture independent surveys andassociated to the class Syndinea (order Syndiniales). These are divided intotwo main groups, MALV-I and MALV-II, the phylogenetic placement ofwhich is still uncertain (Massana & Pedros-Alio, 2008). Both groups have





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been proposed to correspond to heterotrophic parasites of marine organisms(Guillou et al., 2008; Gunderson, John, Boman, & Coats, 2002).

p0185 Dinoflagellates and Apicomplexa are the only groups of alveolates thatpossess plastids. These structures are actively involved in photosynthesis onlyin dinoflagellates, while they are vestigial in apicomplexans (Mcfadden,Reith, Munholland, 1996). This common character between these twogroups suggests that their common ancestor was photosynthetic (Leander &Keeling, 2003). In contrast to other alveolates, dinoflagellates can developcellulose-like polysaccharide plates within the cortical alveoli, forminga theca. The arrangement and ornamentation of these plates leads to anastonishing variety of shapes, the description and comparison of which formsthe traditional basis of species classification for dinoflagellates.

s0095 4.2. Dinoflagellate Diversity

p0190 The majority of dinoflagellates (perhaps 80%) are free-living marineplanktonic or benthic flagellates, the remainder inhabiting equivalentfreshwater habitats (Taylor et al., 2008). Half of the estimated 2000 extantspecies of dinoflagellates are considered photosynthetic. Dinoflagellatescontain a characteristic nucleus with permanently condensed chromosomes(the dinokaryon). The haploid motile stage within dinoflagellate life cyclestypically possesses two dissimilar flagella: a ribbon-like flagellum withmultiple waves situated in a transverse groove (cingulum) and a moreconventional flagellum emerging from a ventral furrow (sulcus) which beatsposteriorly to the cell. After sexual recombination, dinoflagellates produceresistant diploid benthic stages (hypnozygotes, also termed resting cysts) toescape predation and adverse environmental conditions as well as to colonizeecosystems. Some dinoflagellates are noxious to humans and other marineorganisms due to the production of potent toxins, while others causehypoxia, anoxia or mechanical damage to marine fauna. Finally, certaindinoflagellates, including some toxin-producing species, can form largeblooms (‘red tides’) that can negatively affect economic activities in coastalareas (Hallegraeff, 2003, 2010) (Fig. 1.4).

p0195 Nine major orders (Gonyaulacales, Peridiniales, Gymnodiniales, Sues-siales, Prorocentrales, Dinophysiales, Blastodiniales, Phytodiniales andNoctilucales) are recognized within the dinoflagellates (Fig. 1.4), the orderThoracosphaerales being suspected to belong to the Peridiniales (Saldarriagaet al., 2004). Dinoflagellate orders can be distinguished on the basis of majormorphological characters and life cycle features of their members





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(Table 1.1). The Gonyaulacales and Peridiniales are characterized by thepresence of cellulose-like thecal plates within the cortical alveoli. The thecaeof both groups are constituted of five latitudinal series of plates (apical,anterior intercalary, precingular, postcingular and antapical) plus the cingularand sulcal series. The two orders were separated by Taylor (1980). Membersof the order Blastodiniales have a parasitic lifestyle and a temporary dino-karyon, which fostered the hypothesis that Blastodiniales diverged early

t0010

Table 1.1 Main Morphological Features for Major Orders of Dinoflagellates

Order Main morphological featuresGonyaulacales Cellulose-like thecal plates within the cortical alveoli

Left-handed torsion of the epithecaSmall anterior intercalary plates (often absent)Apical pore complex often with a hook-like grooveAsymmetric antapical plates

Peridiniales Cellulose-like thecal plates within the cortical alveoliBilateral symmetryAnterior intercalary platesSmall apical poreTwo subequal antapical plates

Gymnodiniales Cortical alveoli without thecal plates (athecate, unarmored,naked dinoflagellates)

Apical furrow on the cell apexSuessiales Cortical alveoli without thecal plates (athecate, unarmored,

naked dinoflagellates)Generally 7e10 longitudinal series of cortical alveoli (lessthan in Gymnodiniales)

Elongated apical vesicle on the cell apexProrocentrales Division of theca into lateral halves joined by a sagittal suture

Lack of the sulcus and the cingulumTwo pores, the flagella emerging from the larger oneTiny periflagellar platelets

Dinophysiales Division of theca into lateral halves joined by a sagittal sutureTwo pores, the flagella emerging from the larger one

Blastodiniales Temporary dinokaryonParasitic lifestyleDinospores with peridinioid plate tabulation

Phytodiniales Shift from a non-calcareous coccoid cell or continuous-walled colonial stage to a vegetative stage

Noctilucales Highly mobile ventral tentacleLack (at least in some life stages) of the ribbon-like transverseflagellum or the condensed chromosomes of thedinokaryon





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from the dynokaryotic dinoflagellate lineage (Saldarriaga et al., 2004).However, dinospores of the genus Blastodinium have been shown to haveperidinioid plate tabulation, suggesting affiliation to the Peridiniales.Although the genus Blastodinium has been demonstrated to be late-branching, consistent with Peridiniales evolution, the affiliation of the genusand of the whole order of Blastodiniales to Peridiniales is still to be proved(Skovgaard, Massana, & Saiz, 2007). The Prorocentrales and the Dino-physiales share a major synapomorphic feature, unique within dinoflagel-lates: the division of theca into lateral halves joined by a sagittal suture. Theorder Phytodiniales includes species characterized by a shift from a non-calcareous coccoid cell or continuous-walled colonial stage to a vegetativestage. Similar life shifts have, however, also been observed in genera of otherorders (e.g. Suessiales [Symbiodinium], Gonyaulacales [Pyrocystis]). ThePhytodiniales presently includes poorly understood genera for which littlemolecular data are available. The Noctilucales is an early-diverging orderthat includes aberrant dinoflagellates characterized by a highly mobileventral tentacle, which is missing in typical dinoflagellates and other alve-olates. The tentacle does not play a role in keeping the cell in suspension butseems rather related to food capture. The Noctilucales have the ability toincorporate, replace or lose chloroplasts, a rare phenomenon in otheralveolate groups. Whether these chloroplasts are kleptoplastids or derivefrom ancient endosymbiosis, as in other dinoflagellate families, remain to bedemonstrated (Gomez, Moreira, & Lopez-Garcia, 2010).

s0100 4.3. Dinoflagellate Evolution

p0200 To date, comparison of morphological, cytological and nuclear geneticmarkers (18S rRNA gene, 28S rRNA gene, ITS rDNA) has not clearlyresolved relationships between dinoflagellate orders. This raises the questionas to whether all dinoflagellate orders emerged about the same time duringa major radiation period (Hoppenrath & Leander, 2010). Phylogeneticstudies carried out using plastid genes (rbcl, atpB) are limited since manydinoflagellates are heterotrophic, and mitochondrial genes (cob and cox 1) areonly useful in combination with other genetic markers and for specificgroups of dinoflagellates (Zhang, Bhattacharya, & Lin, 2005). Some majorphylogenetic traits can, however, be identified for dinoflagellates (Fig. 1.4).The genus Oxyrrhis is often considered a predinoflagellate lineage(Saldarriaga et al., 2003), followed byNoctiluca scintillans (order Noctilucales),which now appears more closely related to Syndinea (Gomez, Moreira, &





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Lopez-Garcia, 2010). The Dinophysiales (Gomez, Lopez-Garcia, &Moreira, 2011), Suessiales (Siano, Montresor, Probert, Not, & De Vargas,2010) and Gonyaulacales (Saldarriaga et al., 2004) are strongly supportedholophyletic groups. The Prorocentrales splits into two branches in both18S rRNA and 28S rRNA genes phylogenies (Saldarriaga et al., 2004), butnot in mitochondrial cox-1 topology (Murray, Ip, Moore, Nagahama, &Fukuyo, 2009). Cox-1 demonstrates the monophyly of the morphologicallyvery cohesive group of prorocentroid dinoflagellates, but it separates thegroup into two clades that include species present, respectively, in the twobranches of the 28S rRNA gene phylogeny. The Gymnodiniales is a poly-phyletic order and together with the Peridiniales are the most evolutionarycomplex groups of dinoflagellates. The Karenia/Karlodinium clade separatesfrom other gymnodinioid clades (e.g. Akashiwo, Gymnodinium, Amphidiniumclades) in both 18S and 28S rRNA gene phylogenies (Murray, Jorgensen,Ho, Patterson, & Jermilin, 2005; Saldarriaga et al., 2004). The Peridinialesappears to be a complex paraphyletic group of dinoflagellates, that is, itsphylogeny comprises non-Peridiniales branches. The Heterocapsa clade oftenhas a basal position to other peridinioids and in general to other thecatedinoflagellates in phylogenies inferred from the 18S and 28S rRNA genes,despite having mediocre branching support. Other Peridiniales branches(Peridinium, Scrippsiella and Protoperidium) branch later in phylogenetic trees.General dinoflagellate phylogenies still require molecular data for manydinoflagellate genera (especially for heterotrophic species), taxonomicrevision of some species and identification of species likely to correspond tomissing branches of phylogenetic trees (Fig. 1.4).

p0205 Considerable diversity of chloroplast types and pigment compositionoccurs in photosynthetic dinoflagellates, acquired through secondary andtertiary endosymbioses (Cavalier-Smith, 1999) with in some cases multiplelosses and replacement of plastids, as revealed by molecular phylogeneticanalysis (Saldarriaga et al., 2001). Some dinoflagellates harbor foreign plastidsthat are periodically lost and gained during their life cycle (kleptoplastidy,from the Greek ‘klepto’ – stealing), or bear photosynthetic endosymbiontsthat are kept for longer periods but not fully integrated (Moestrup &Daugbjerg, 2007). Beyond chlorophyll a, c2 and b-carotene, dinoflagellateswith permanent plastids can have three other accessory pigments: perdinin(Gonyaulacales, Peridiniales, Prorocentrales, Suessiales and some Gymno-diniales), fucoxanthin or fucoxanthin derivatives (the family Kareniaceaeof the order Gymnodiniales) and chlorophyll b (the genus Lepidodiniumof the order Gymnodiniales). The toxin-producing genus Dinophysis





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(Dinophysiales) is a peculiar case. It is still debated whether Dinophysis acu-minata has permanent plastids of cryptophyte origin (Garcia-Cuetos,Moestrup, Hansen, & Daugbjerg, 2010) or whether it maintains a temporaryplastid (kleptoplast) acquired from prey (Wisecaver & Hackett, 2010). Thelatter hypothesis is supported by the fact that Dinophysis species can only becultured using the ciliate Myrionecta rubra as prey, which itself feeds oncryptophytes and would thus be the source for Dinophysis of the crypto-phytes and their chloroplasts (Park et al., 2006). The fact that Dinophysisneeds the ciliate as an intermediary to acquire cryptophyte plastids suggeststhat it probably lacks critical enzymes to initially process the prey or maintaintheir plastids (Wisecaver & Hackett, 2010). Future transcriptomic andgenomic sequencing would help up to understand if this dinoflagellatepossesses a photosynthetic machinery and how it regulates genes thatcoordinate photosynthetic activity and/or prey capture.

s0105 4.4. Ecology of Dinoflagellates

p0210 Photosynthetic dinoflagellates are common and abundant in pelagic andbenthic habitats of both marine and freshwater ecosystems. Typically, theyreach their highest abundances in estuaries and coastal marine waters, inconcomitance with high nutrient supply from land sources and/or deepwater upwelling. Blooms of noxious species (HABs) are more commonunder these conditions (Fig. 1.4). Using their two perpendicular flagella,dinoflagellates exhibit directed movement in response to chemical stimuli,physical variations, gravity and light. Due to this motility, dinoflagellates areable to find optimal conditions for growth and survival under high physicaldisturbance (turbulence and shear forces), intense light stress and nutrientlimitation. Because dinoflagellates have various habitat preferences, multiplelife strategies and are nutritionally versatile, they are very competitive withother groups of protists for resource acquisition. Although they can some-times be important in terms of biomass, both micro- and nanodinoflagellatesare rarely reported to dominate in terms of abundance within the photo-trophic fraction of plankton communities (Siokou-Frangou et al., 2010).This is probably partly due to the fact that quantitative information is veryfragmentary for dinoflagellates because of inadequate identification meth-odologies, notably for athecate species.

p0215 In coastal waters, bloom initiation can be due to germination of vege-tative cells from hypnozygotes. The biological and physical factors thattrigger bloom initiation are poorly known for most dinoflagellates, including





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harmful species (Burkholder, Azanza, & Sako, 2006). During bloomdevelopment, many dinoflagellate species are capable of rapid growth,attaining abundances up to 109 cells/L (up to 400–500 mg Chla/L) (Taylor &Pollingher, 1987). Dinoflagellates can grow at rates of up to 3.5 divisions perday, but only 15% of the larger free-living harmful species have growth ratesgreater than 1.0 division per day (Smayda, 1997). Photosynthetic dinofla-gellates are primarily limited by phosphorous and nitrogen, although theycan store these nutrients in a species-specific way that in some cases allowsone species to outcompete others (Graham & Wilcox, 2000; Labry et al.,2008). As for other phytoplankton taxa, micronutrients, including forms ofselenium and iron, have been shown to influence blooms of some harmfulphototrophic dinoflagellates (Boyer & Brand, 1998; Doblin, Blackburn, &Hallegraeff, 2000). Apart from nutrient limitation, bloom termination canbe caused by water dispersion and dilution, zooplankton grazing and bio-logical endogenous cycles, as well as parasite (Chambouvet, Morin, Marie, &Guillou, 2008) and viral (Nagasaki, Tomaru, Shirai, Takao, & Mizumoto,2006) infections.

p0220 Field observations indicate that bloom-forming dinoflagellate specieshave neither strict habitat preferences nor uniform responses. Smayda andReynolds (2001) recognized nine different pelagic habitats where dinofla-gellates bloom, arranged along an onshore–offshore gradient of decreasingnutrients, reduced mixing and deepening of the photic water layer. Each ofthe nine types of habitat is characterized by a specific dinoflagellate life-form,which suggests that dinoflagellates have evolved multiple adaptive strat-egies,rather than a common ecological strategy. Subsequently, these authorsintroduced five rules of assembly for marine dinoflagellate communities,which state that specific habitat conditions correspond to specific life formsthat are mainly selected on the basis of abiotic factors (turbulence andnutrient availability). Within the species pool of a given habitat, seasonalsuccession is stochastic and characterized by a high degree of unpredictability(Smayda & Reynolds, 2003). Conversely, for cyst-forming dinoflagellates,the existence of endogenous or exogenous factors determining the presenceand succession of species has been hypothesized, suggesting that the apparentrandom succession of species within a pool of species is understandable andpredictable (Anderson & Rengefors, 2006).

p0225 The ecological role of dinoflagellates in the functioning of marineecosystems and in the marine food web can be significant. Some hetero-trophic species of the genera Ornithocercus, Histioneis and Citharistes can hostphotosynthetic endosymbionts. In autumn in oligotrophic subtropical





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waters of the Gulf of Aqaba (Israel), peaks of these species coincided withextended nitrogen limitation and high abundances of free-living N-fixingcyanobacteria. It has been proposed that heterotrophic dinoflagellate hostsmay provide cyanobacterial symbionts with the anaerobic microenviron-ment necessary for efficient N fixation, which would in turn determine theecological success of the hosts (Gordon, Angel, Neori, Kress, & Kimor,1994). This symbiotic relationship as well as the capacity of N fixation bythe cyanobacterial symbionts has been suggested (Foster, Carpenter, &Bergman, 2006). Alternatively, some dinoflagellates are symbionts ofbenthic organisms (Symbiodinium) (Freudenthal, 1962) or of pelagic protists(Pelagodinium) (Siano et al., 2010). Many, if not most, photosyntheticdinoflagellates are considered mixotrophic (Smalley & Coats, 2002). It hasbeen suggested that the larger size and lower cell surface-to-volume ratios ofdinoflagellates generally result in lower affinities for dissolved nutrients thansmaller protists, therefore positively selecting for mixotrophy amongphotosynthetic species (Smayda, 1997). However, mixotrophy is oftendifficult to assess clearly because of low feeding rates, intermittent feedingdependent on conditions poorly simulated in cultures, specificity of prey orthe fact that organelles can obscure food vacuoles (Stoecker, 1999).

s0110 5. THE SILICEOUS PHYTOPLANKTON: THE DIATOMS

p0230 Diatoms, also called Bacillariophyceae, are unicellular photoautotro-phic stramenopiles, the defining feature of which is the compound silica cellwall, called a frustule. The diatoms constitute one of the most diverselineages of eukaryotes with possibly over 100,000 extant species (Mann &Droop, 1996). They are ubiquitous in marine and freshwater habitats and indamp terrestrial environments. It is therefore not surprising that diatomshave been studied intensively ever since microscopes became available. Wereview here the diversity and ecology of diatoms and refer to Chapter VII ofthis volume for a review on diatom genomics (Mock & Medlin 2012).

s0115 5.1. The Hallmark of the Diatom: The Silica Cell Wall

p0235 The diatom frustule is composed of two overlapping thecae (the larger calledthe epitheca and the smaller the hypotheca), each of which consists of a valveand an accompanying series of girdle bands. These frustule elements containrows of pores, called interstriae, with ribs between them, called striae. Thepores in the interstriae form the principal conduits for the uptake of nutrients





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and exudation of metabolites. Girdle bands are architecturally relativelysimple and always consist of a single layer, whereas valves are more elaborate,with a flat area, called the valve face, and a rim, called the mantle. Valves areeither composed of a single layer or two layers. In the latter case, the internallayer has larger pores and the two layers are connected via perpendicularcross walls in a rectangular or honeycomb pattern, giving rise to chamberedvalves (see Round, Crawford, & Mann, 1990).

p0240 Construction of new silica cell wall elements proceeds in silica depositionvesicles (SDVs) near the plasma membrane. Dissolved silicic acid is activelytaken up from water and concentrated in the cytoplasm far above the level atwhich silica would normally polymerize. Precipitation in the cytoplasm isavoided because silicic acid transporter proteins bind the silica and shepherdit through the cytoplasm into the SDV. Other classes of peptides, includingsilaffins, silacidins and long-chain polyamines, are produced in the endo-plasmic reticulum, transported in vesicles to the Golgi apparatus where theyare modified and activated, then transported into the SDV where they forma matrix onto which the supersaturated silica precipitates in an amorphousform (Hildebrand, 2008; Kroger & Poulsen, 2008).

p0245 During vegetative cell division, new thecae are laid down in such a waythat the cell is covered completely by silica cell wall elements throughout thedivision phase. Newly formed thecae must therefore be formed within theconfines of existing thecae. The daughter cell inheriting the parental epi-theca (the larger half of the frustule) forms a new hypotheca of the same sizeas that of the parental cell and hence this daughter cell is of the same size asthe parent. By contrast, the daughter cell inheriting the parental hypotheca(the smaller half of the frustule) uses it as an epitheca within which a newhypotheca is formed; hence, this daughter cell is slightly smaller than itsparent. Consequently, average cell diameter diminishes with ongoingmitotic division. The only escape from this continual miniaturization issexual reproduction (see Section C).

s0120 5.2. Diatom Diversity

p0250 Diatoms are categorized into centrics and pennates based on characteristics ofvalves, including shape, ultrastructure, ornamentation and types of processes(tubes, slits) (Fig. 1.5). The principal difference is the way in which theinterstriae are organized. Centric diatoms possess radially organized valveswith striae radiating from a central region or ring, whereas pennates possesselongated valves with striae oriented perpendicular to a midrib (called





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a sternum), like in a feather (Round et al., 1990). Electron microscopeillustrations of ultrastructural details of diatoms can be found in Round et al.

p0255 One group of centric diatoms, the radial centrics, possesses valves shapedlike Petri dishes. Radial centrics usually possess a ring of so-called labiateprocesses around their valve mantle, forming tubes that may enable intake orsecretion of organic material. Important planktonic representatives of radialcentrics include Aulacoseira, Corethron, Coscinodiscus, Leptocylindrus, Melosiraand Rhizosolenia. A second group of centrics, the so-called multipolarcentrics (bi-, tri-, etc., referring to polarity of shape) also exhibit a radial poreorganization, but their cell form is usually elongate, triangular or starlike;that is, exhibits polarity. Labiate processes, if present, are located on thecentral area of valves. Many species possess fields of densely packed pores attheir valve apices. Mucilage is exuded through these apical pore fields,enabling benthic or epiphytic species to attach to the substratum or to formchains. Planktonic representatives such as Eucampia also form chains this way.Secondarily radial centric genera such as Lauderia, Porosira, Skeletonema andThalassiosira (order Thalassiosirales) possess specialized tubes in their valves,called strutted processes, through which chitin filaments are exuded. Thesefilaments extend into the surrounding medium and link cells into chains.Chaetoceros and Bacteriastrum (order Chaetocerotales) form chains by meansof setae; thin hollow silica tubes that are formed following cell division.Setae probably evolved principally as a grazer deterrent, but they haveacquired additional functions. In some species, plastids migrate up and downsetae. Chaetocerotales and Thalassiosirales are highly diverse and theirspecies form important constituents of phytoplankton blooms.

p0260 Pennate diatoms are elongate, their defining feature being the midribfrom which striae and interstriae extend perpendicularly. The pennates aresubdivided into raphid pennates, which possess a slit-shaped process, calleda raphe, and araphid pennates, which lack a raphe, but have apical pore fieldsand apical labiate processes. The raphe slit enables raphid pennates to moveactively by means of a cost-effective system of traction. Long-chain organiccompounds protrude partially through one end of the slit and connect to thesubstratum. Subsequently, the chain is pulled along the slit and then exuded.Consequently, the diatom moves in the opposite direction. Most raphidpennates are benthic, but a few, including Fragillariopsis and Pseudonitzschia,have secondarily acquired a planktonic lifestyle. The araphid pennates arealso typically benthic, but there are also several lineages that have acquireda planktonic lifestyle, including Asterionella, Asterionellopsis, Lioloma, Tha-lassionema and Thalassiothrix.





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p0265 Arguably most diatom diversity is not planktonic, but benthic orepiphytic. Many species occur unattached, but their frustules are robust andway too heavy for a planktonic existence, thus abounding drifting oversandy bottoms. Diatom diversity shows several clades and grades containingplanktonic species, but these are most often rooted in benthic ancestry. Thisversatility between planktonic and benthic existence in the evolutionaryhistory of the diatoms means that traits acquired in a benthic setting may laterhave provided a benefit in the plankton. For example, raphid pennatediatoms are typically found in benthic habitats where they use their raphe tomove actively over the substratum. Pseudonitzschia, a planktonic represen-tative that evolved from benthic ancestry, use their raphe to enable daughtercells to slide along each other adjacent valve faces to assume a position inwhich they are just attached at their valve apices. In this way, Pseudonitzschiahas acquired a novel way to form chains in the plankton.

s0125 5.3. Diatom Life Cycle

p0270 The diatoms possess a diplontic life cycle consisting of a long period (up toseveral years) during which diploid cells divide mitotically and a brief period(a few days) during which sexual reproduction takes place. Gametes fuse toform a zygote, which inflates to generate an auxospore. The new frustuleelements of the resultant vegetative cell are formed within the confines of theauxospore wall and once this is completed the cell emerges from the aux-ospore (Round et al., 1990). Centric diatoms form non-motile macrogametesand flagellated microgametes. In radial centrics and Thalassiosirales, thezygote inflates isometrically and forms an organic wall in which, in somespecies, small silica elements are embedded. In multipolar centrics, gameteformation and conjugation proceed as in radial centrics, but zygote inflation isanisometric; in this case, a series of silica bands, together called a proper-izonium, is laid down in sequence to mould the expanding auxospore intoa bi- or multipolar shape. Pennate diatoms are isogamous (¼ producegametes of equal size, though generally not of equal behaviour) and usuallydioecious. Sexual reproduction involves alignment of cells of oppositemating types, gamete formation, migration in an amoeboid fashion to thepartner gametes and zygote formation. Zygote inflation is constrained bya double series of perizonial bands into a cigar-like auxospore. The newfrustule is formed within the confines of this perizonium. For pennates, it isunclear what provokes sexual reproduction, how potential mates detect eachother and how and in what order they initiate the process of gametogenesis.





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p0275 Several centric diatoms form resting spores (Ishii, Iwataki, Matsuoka, &Imai, 2011; Kooistra et al., 2010). Spores are formed usually in response todeteriorating conditions. Resting spores can remain viable for extendedperiods (H€arnstr€om, Ellegaard, Andersen, & Godhe, 2011) and are often sorobust that they exist as fossil markers (Suto, 2006). Auxospores are notresting cysts; their silica elements are flimsy and disintegrate and dissolvesoon after the initial cell hatches.

s0130 5.4. Diatom Evolution

p0280 Results of molecular phylogenetic studies have generally revealed thatcentrics are the most ancient group of diatoms, forming a grade (¼ para-phyletic group). Multipolar centrics evolved from radial centric ancestry, butthey form a grade because all pennates form a clade within this group.Within the pennates, the raphid pennates form a clade inside a grade ofaraphid pennates (Fig. 1.5).

p0285 This phylogenetic pattern suggests that the properizonial bands ofmultipolar centric auxospores and the perizonial bands of pennate aux-ospores are homologous structures and that these bands were acquired in thelast common ancestor of the clade containing all multipolar centrics andpennates. The formation of these bands during auxospore formation enableddiatoms to generate shapes diverging from those resembling Petri dishes andtubes. Moreover, the results suggest that isogametogenesis is a derived trait ofall pennates and that it evolved from oogenesis of their multipolar ancestor,possibly because it is a more efficient mode of sexual reproduction in benthicenvironments. The organization of the pennate valve with its midribevolved from a radial organization mode. The advantage of such a midribmay have been the structural reinforcement it provides in lightweight butelongated diatom valves. Finally, the raphe of raphid pennates evolved fromthe apical labiate processes of their araphid pennate ancestry (Kooistra,Gersonde, Medkin, & Mann, 2007).

p0290 Each of these acquisitions apparently resulted in rapid diversification ofthe lineage exhibiting the novelty. Radial centrics appear to consist ofa restricted number of remnant lineages, as do most multipolar centrics.Within the latter, the Thalassiosirales with their chitin threads and theChaetocerotales with their setae constitute two highly diverse clades. Thepennates as a whole form a huge clade with a poorly resolved basal structureand the same is true for raphid pennates that constitute a relatively novellineage, but by far the most diverse group. Nonetheless, radial centrics do not





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constitute an evolutionary dead-end. Many radial centrics, such as Lep-tocylindrus, are important bloom formers in coastal regions all over the world.

p0295 Diatoms possess an extensive fossil record. Frustule elements of vegeta-tive cells and spores are preserved, often in exquisite detail, over millions ofyears (e.g. Gersonde & Hardwood, 1990; Ishii et al., 2011; Suto, 2006). Inaddition, biomarkers specific for particular diatom lineages, for instanceRhizosolenia, are detectable in petroleum of definable age (Sinninghe-Damsté et al., 2004). These sources of information reveal that radial centricsfirst appeared in the Jurassic, multipolar centrics in the Early Cretaceous,pennates in the Late Cretaceous and the first raphid pennates at 55 Mya inthe Paleogene. This order of appearance corroborates the above-mentionedphylogenetic patterns in extant diversity. The diatoms apparently traversedthe K/T boundary relatively unscathed, with diversity subsequentlyincreasing (Harwood, Chang, & Nikolaev, 2004; Stoermer & Smol, 1999).

s0135 5.5. Diatom Ecology

p0300 Diatoms are ubiquitous in the plankton and benthos of marine and fresh-water habitats. Centric diatoms are typically marine, but a few exclusivelyfreshwater genera exist and a few large marine genera have freshwaterrepresentatives (e.g. Thalassiosira). Araphid pennates are predominantlymarine, but there are also many freshwater genera and some marine generahave freshwater representatives. Raphid pennates seem to be well repre-sented in both freshwater and marine environments and many genera haverepresentatives in both. Because of their abundance in shallow coastal seas, ithas been estimated that planktonic diatoms account for as much as 20% ofglobal photosynthetic fixation of carbon (w20 Pg carbon fixed per year;Mann, 1999), which is more than all the world’s tropical rainforests, and thattheir contribution to nutrient cycling is significant (Boyd et al., 2000).Planktonic diatoms are particularly important bloom-formers in nutrient-rich coastal regions, especially in temperate zones and upwelling zones inwarm-temperate and tropical regions. Two characteristics render themparticularly well suited to such environments. Firstly, they are well adaptedto growth in deeply mixed turbulent water, where cells are only intermit-tently exposed to high light levels (Falkowski & Raven, 1997). Secondly,they are well adapted to pulsed availability of nutrients because they can usetheir often large central vacuole for nutrient storage. Diatoms are heavilygrazed upon by a plethora of herbivores, in particular copepods. Diatomsseem to have few viral adversaries, but they are subject to attack from a range





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of protistan parasites such as oomycetes and thraustochytrids (Hanic, Seki-moto, & Bates, 2009). Defense mechanisms against biological pressuresinclude structural reinforcement of frustule elements with double layers andinternal struts and buttresses, such as in the pennate diatom Fragillariopsis andin many radial centric diatoms, setae in Chaetoceros and Bacteriastrum, barbedspines in Corethron, or barbed spine-like extensions of the valve as inAsterionellopsis. Other strategies include biochemical defense by means ofproduction of metabolites that become bioactive when the diatom cell isdisrupted by grazers. Biological attack results in a large part of diatomproduction being rapidly remineralized in surface waters. Nonetheless,a considerable part of the primary production of coastal diatom blooms sinks.The bulk of this organic material is re-mineralized in deeper water, butgiven high sedimentation rates in coastal regions, a small part is trapped insediments. In this way, coastal planktonic diatoms contribute to about half oftotal long-term organic carbon sequestration in the marine environment.This is why petroleum is found in present and past river plume sedimentsalong so-called passive continental fringes (Brazil, the North Sea, CaspianSea, Venezuela, Gulf of Mexico and the Middle East).

p0305 Diatoms are also important constituents of phytoplankton communitiesin the Southern ocean. In this region, diatom cell densities are typically low,mainly because of restricted availability of iron, but population sizes arenevertheless considerable given the extent of the region. Southern oceandiatoms possess well-developed physical grazer defenses in the form of spines(e.g. Corethron, Chaetoceros) or chambered and structurally reinforced valveelements (e.g. Fragillariopsis). Diatoms also abound (but do not dominate) inthe deep chlorophyll maximum of warm open oceans. In these conditions,species are generally minute and do not form chains, but their diversity ispoorly known; hence, it is here that molecular taxonomic surveys mightuncover considerable new diversity. Diatoms do not contribute significantlyto long-term carbon sequestration in any of these open oceanic communities.

s0140 6. LAST, BUT NOT LEAST RELEVANT: OTHERPHYTOPLANKTON TAXA

s0145 6.1. Stramenopiles Other Than Diatoms

p0310 Within the stramenopile lineage, a number of other phytoplankton groupsexist besides diatoms (Fig. 1.1). Although less diverse, some of these groupshave important ecological roles in marine ecosystems. They are usually





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flagellated cells with heterokont characteristics, that is, two unequal flagella,one being ornamented with hair-like structures called mastigonemes. Theypossess plastids acquired through secondary endosymbiosis, typically withchlorophylls a and c. Their main characteristics are summarized in Table 1.2.

t0015 Table 1.2 Main Characteristics of Stramenopiles Containing PhytoplanktonicRepresentatives Other Than Diatoms

Taxonomy Main characteristicsBolidophyceae Exclusively marine picoplankton

Fast swimming cells (Guillou et al., 1999)Phylogenetically linked to Parmales (i.e. non-motile cells covered with silica plates thatare anisometric with a radial organizationsimilar to that of the valves of centricdiatoms (Ichinomiya et al., 2011)

Dictyochophyceae (alsocalled silicoflagellates byprotistologists)

Characterized by tentacules or rhizopodiaPresence of a siliceous skeleton in onephase of the life cycle in the orderDictyocales

Pelagophyceae No real distinct morphological character butsupported as a disctinct lineage in allphylogenies

The order Pelagomonadales contains mainlypicoplanktonic representatives (e.g.Pelagomonas, Aureococcus), while the orderSarcinochrysidales contains bothplanktonic and benthic genera

Raphidophyceae Characterised by the presence of ejectilebodies (trichocysts)

Cells with two flagella, without cell wallPresence of mucilaginous bodies underplasmalemma in some genera

Marine raphidophytes form a monophyleticclade and have a pigment compositiondistinct from their freshwater counterparts(Yamaguchi, Nakayama, Murakami, &Inouye, 2010)

Pinguiophyceae No real distinct morphological character butsupported as a disctinct lineage in allphylogenies

Characterised by the production of largeamounts of omega-3 fatty acids(polyunsaturated fatty acids)





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Distinctive derived characters (synapomorphies) for each class are indicatedwhen possible. However, it is sometimes difficult to define classes usingmorphological characters and a combination of ultrastructural andbiochemical features is often needed, while some classes are only definedbased on phylogenetic analyses.

p0315 The stramenopile group that is phylogenetically nearest to diatoms isthe Bolidophyceae (Table 1.2). Bolidophyceae are regularly detected inmolecular surveys and isolated in to culture from the marine environ-ment, but very few quantitative studies have been performed and theirecological imprint is still unclear (Guillou, 2011). The dictyochophytesare frequently observed in the environment and are regularly detected inmolecular surveys of planktonic communities (Massana & Pedros-Alio,2008). Several species have been recently described (e.g. Florenciellaparvula, Eikrem, Romari, Gall, Latasa, & Vaulot, 2004) or renamed andtransferred from the raphidophyte genus Chattonella with whichconfusions were frequent (e.g. Pseudochattonella sp., Hosoi-Tanabeet al., 2007). Some dictyochophyte species can produce icthyotoxins(Skjelbred, Horsberg, Tollefsen, Andersen, & Edvardsen, 2011). Pela-gophytes are essentially known because of the ecosystem disruptive algalblooms of the brown tide species Aureococcus anophagefferens and Aur-eoumbra lagunensis (Gobler & Sunda, 2012). Other species such as Pela-gomonas or Pelagococcus are frequently isolated from sea water, and theirabundance, estimated through pigment signatures, indicate that they areprobably non-negligible phytoplankton contributors in oligotrophic

Table 1.2 Main Characteristics of Stramenopiles Containing PhytoplanktonicRepresentatives Other Than Diatomsdcont'd

Chrysophyceae/Synurophyceae Taxa in both classes characterised by theability to form silicified cysts or statospores

Most species in these classes inhabitfreshwater. Taxa covered by siliceous scalesin both classes

The Chrysophyceae have been re-definedrecently (Andersen, 2004)

Eustigmatophyceae Characterised by the absence of chlorophyll c(lost through evolution)

Zoospores elongated with a basal swelling onthe anterior flagellum. Stigma is incytoplasm (and not in plastid)





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regions (Not et al., 2008; Shi et al., 2011). The Pinguiophyceae is a classof small-sized phytoplankton erected in 2002 (Kawachi et al., 2002).One of their distinctive features is the production of large amounts ofomega-3 fatty acids (polyunsaturated fatty acids), providing them witha yet unexploited potential for biotechnological applications (Kawachiet al., 2002). They are rarely found in environmental surveys and do notseem be major contributors to phytoplanktonic communities (Fulleret al., 2006).

p0320 The raphidophytes, chrysophytes, synurophytes and eustigmatophyteshave marine representatives but are most abundant and diverse in fresh-water. Yet, some species are of primary importance in marine ecosystems.Raphidophytes are found worldwide, mostly in coastal regions. The generaHeterosigma and Chatonella can cause important harmful effects in coastalwaters and thus have a significant economic impact on aquaculture (Imai &Yamaguchi, 2012). Plastidial and mitochondrial genomes have recentlybeen sequenced and analyzed (Masuda et al., 2011). The most commonmarine eustigmatophyte genus is Nannochloropsis, a small-sized phyto-plankton (2–4 mm) for which six species are currently described (Andersen,Brett, Potter, & Sexton, 1998). They have a characteristic pigmentsignature with violaxanthin and vaucheriaxanthin like as dominatingcarotenoids (Karlson, Potter, Kuylenstierna, & Andersen, 1996) and areessentially studied because they produce large quantities of fatty acids andin particular omega 3. Strains of Nannochloropsis are commonly used inaquaculture and because these microalgae are good candidates for greenenergy and blue biotechnologies (Cadoret, Garnier, & Saint-Jean, 2012),Nannochloropsis is one of the few phytoplankton taxa for which a fullgenome sequence is available (Kehou et al., 2011; Oliver, Benemann,Niyogi, & Vick, 2011). Although regularly found in the environment,relatively few studies have been performed on the ecology of eustigma-tophytes. Chrysophytes and synurophytes are closely related phylogenet-ically. Chrysophytes can be strictly phototrophs or heterotrophs or presenta dual mixotrophic strategy (Preisig, Vørs, & H€allfors, 1991). A typicalgenus is Ochromonas comprising about 80 known species found in bothfreshwater and marine environments. Recent environmental molecularsurveys of plankton diversity demonstrate the presence of novel clades ofmarine chrysophytes, in particular in the picoplankton size range, with nocultured representatives (Del Campo & Massana, 2011; Fuller et al., 2006;Shi et al., 2009).





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s0150 6.2. The Cryptophytes

p0325 Cryptophytes are unicellular algae characterized by features including (1)asymmetrical cell shape, (2) presence of an invagination (either a tubulargullet, a furrow, a combination of furrow and gullet or a groove) lined withstructures termed ejectosomes that discharge ribbon-like threads uponmechanical or chemical stress (Morrall & Greenwood, 1980), (3) a layeredstructure surrounding the cells that consists of proteinaceous inner andsurface periplast components sandwiching the plasma membrane (Brett,Perasso, & Wetherbee, 1994), (4) a plastid surrounded by four membranes,with the periplastidial space containing a highly reduced remnant nucleustermed the nucleomorph (Gillott & Gibbs, 1980) and (5) a light-harvestingcomplex consisting of chlorophylls a and c2, xanthophylls, and either red orblue phycobiliproteins (Hill & Rowan, 1989). Different combinations and/or concentrations of the pigments give cryptophytes brown, red or blue-green coloration. Cryptophyte cells have two unequal flagella, the longerone with two opposite rows of stiff flagellar hairs that provide reverse thrust,the shorter one with a single row of flagellar hairs (Hibberd, Greenwood, &Griffiths, 1971). The flagella are inserted subapically in the vestibule of aninvagination often shifted to the cell’s right, whereas the cell apex is shiftedto the left.

p0330 Most cryptophytes are photosynthetic, but some have lost their photo-synthetic pigments and returned to a heterotrophic mode of nutrition withretention of a leucoplast (remnant chloroplast) (Hoef-Emden, 2005). Thephagotrophic Goniomonas is the only known cryptophyte genus withouta plastid (Mcfadden, Gilson, & Hill, 1994) and also differs from plastid-containing cryptophytes in cell shape (flattened in lateral plane), cellinvagination(s) and flagellar structure. Ultrastructural characters such asdifferences in periplast structure, type of invagination, flagellar apparatusand/or position of the nucleomorph have been used to define cryptophytegenera. In addition, each taxon produces only one of the seven known typesof biliprotein. Cryptophytes thrive in all kinds of aqueous habitats, includingmarine, brackish and freshwater, but seem to be more diverse in marine thanin freshwater habitats (Clay, Kugrens, & Lee, 1999; Novarino, 2003).Marine as well as freshwater cryptophytes are often abundant components ofplanktonic communities and may form blooms but are not known to haveharmful impacts. Red-coloured genera like Rhodomonas and Rhinomonas arepredominately marine, blue-green genera such as Chroomonas are present inboth marine and freshwater habitats, while the brown to olive coloured





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genus Cryptomonas is restricted to freshwater habitats (Hoef-Emden &Melkonian, 2003). The origins and phylogenetic affiliations of cryptophytesare uncertain (see haptophyte section above).

s0155 6.3. The Chlorarachniophyta

p0335 Chorarachniophyta are a small group of algae, the first describedmembers of which were amoeboid and associated with debris (Ishida,Yabuki, & Ota, 2007). They present a strong phylogenetic interestbecause they possess a four membrane plastid containing green algalpigments (chlorophyll b in particular) as well as a remnant of theendosymbiont nucleus called the nucleomorph, the genome of which hasbeen sequenced for at least one species (Gilson et al., 2006). At present,only 13 species and 8 genera are known. In recent years, planktonicchlororachniophytes have been described, one of picoplanktonic size(Moestrup & Sengco, 2001; Ota, Vaulot, Le Gall, Yabuki, & Ishida,2009). Their ecological importance remains unknown, although 18SrRNA gene environmental sequences related to chlororachniophyteshave been obtained from the Mediterranean Sea and the upwelling offChile using specific primers (Ota, personal communication). Moreover,several new species have been isolated in to culture from the Mediter-ranean Sea, hinting that this group could have a narrow biogeographicaldistribution (Ota & Vaulot, 2012).

s0160 6.4. The Euglenids

p0340 The Euglenozoa form a monophyletic lineage within the Excavata, a lineagethat contains parasites, photo-autotrophs and predators. Typically, excavateshave flagella inserted into a reservoir, paramylon (a-1,3 glucan) as the maincarbohydrate reserve and a peculiar type of closed mitosis. The photosyn-thetic euglenids, the class Euglenophyceae, acquired a green plastid throughsecondary endosymbiosis (Marin, 2004). They mostly occur in freshwaterhabitats, but a few marine species belonging to the Eutreptiellales areknown.

s0165 7. CONCLUDING REMARKS

p0345 Analysis of the diversity and ecology of phytoplankton has largelybenefited from molecular data, and, as for many research fields in biology,phytoplankton research is entering a new era with the advent of





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high throughput sequencing technologies. Large-scale environmentalmeta-barcoding allows quasi-exhaustive analysis of the diversity ofcommunities. Working at the community level allows integration offunctional information, even for uncultured taxa (Toulza et al., 2012, inthis volume). In the near future, if carefully contextualized with envi-ronmental meta-data, the panel of ‘-omics’ tools now available will facil-itate eco-systemic approaches (Raes & Bork, 2008) to the analysis ofphytoplankton communities and lead to a better understanding ofphytoplankton ecology. It will also promote detailed experimental andphysiological studies on particular taxa, ultimately fostering improvementof predictive models of phytoplankton distribution at global scales (Barton,Dutkiewicz, Flierl, Bragg, & Follows, 2010).

p0350 However, in this environmental genomic context, the improvement ofknowledge on the diversity and ecology of phytoplankton strongly relies onhigh-quality reference databases and is tightly linked to our ability to clearlybridge molecular data and phenotypes. The potential to simultaneously andquantitatively assess the occurrence of phytoplankton taxa at relevant spatialand temporal resolution is also essential. These goals can only be achievedthrough multidisciplinary research involving strong taxonomic expertiseand the development of high throughput microscopy and automaticmolecular and imaging tools (Olson & Sosik, 2007; Preston et al., 2011).While some phytoplankton species can be harmful for ecosystems and/orhuman activities, others provide great potential for providing naturalproducts through blue biotechnology and alternative green energy(Larkum, Ross, Kruse, & Hankamer, 2011). Although little genomic dataare currently available for phytoplankton (Rynearson & Palenik, 2011),genomics and postgenomics approaches will undoubtedly significantlycontribute to unraveling the nature and implications of biological processesacross ecological scales and will help addressing some of the currentconceptual challenges in phytoplankton research and more generally inmicrobial ecology.

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c0001 diversity and ecology of eukaryotic marine phytoplankton...

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